1. Field of the Invention
The present invention relates to the construction of expression cassettes for use in gene expression studies in both homologous and heterologous Streptomyces strain background. Specifically this invention describes construction of two Streptomyces lividans-recombinant strains capable of producing 31-O-desmethylFK-506 O:methyltransferase, which methylales 31 -O-desmethylFK-506 to FK -506.
2. Brief Description of Disclosures in the Art
The economic potential of DNA recombinant technology in producing unlimited amount of the pharmaceutically valuable peptides and proteins is vast. For example, the biologically active peptides and proteins such as insulin, erythropoietin, granulocyte-colony stimulating factor (G-CSF) and other important growth factors can now be produced in an unlimited quantities in bacteria, yeast and mammalian cell lines. Similarly, unlimited amounts of catalytic proteins (enzymes) may also be produced. This is significant because the use of enzymes as catalytic reagents in the synthesis of organic compounds are becoming increasingly practical and popular, not only because of economic factors, but also due to convenience in carrying out the chemical reactions, and environmental concerns where the use of toxic organic chemicals and solvents needs to be reduced or eliminated. In addition, enzymes are capable of catalyzing certain reactions that might otherwise be impossible or difficult to carry out by traditional organic reagents. To exploit DNA recombinant technology, however, it is essential that a large scale fermentation technology is available. Furthermore, it is equally critical that the bioconverting cultures and/or purified catalytic proteins capable of catalyzing particular organic reactions are accessible.
Regarding large scale fermentation, the technology is well developed for the microorganisms belonging to the genus Streptomyces; a large number of the secondary metabolites and extracellular enzymes are commercially produced by the fermentation of this microorganism. With regard to the availability of the bioconverting cultures and/or purified proteins, these cultures and proteins have to be discovered, isolated and/or made by using a genetic engineering approach. There is a long felt need in the art for different cultures which are capable of catalyzing a variety of organic reactions. The present invention meets this need by providing expression systems for the introduction of a single gene into the Streptomyces chromosome which enables the host organism to produce proteins with particular desirable catalytic properties. Furthermore, the present expression systems may be transferred as cassettes onto low as well as high copy Streptomyces replicating plasmids for the production of large quantities of a desired compound.